Controlling antifungal activity with light: Optical regulation of fungal ergosterol biosynthetic pathway with photo-responsive CYP51 inhibitors

Invasive fungal infections (IFIs) have been associated with high mortality, highlighting the urgent need for developing novel antifungal strategies. Herein the first light-responsive antifungal agents were designed by optical control of fungal ergosterol biosynthesis pathway with photocaged triazole lanosterol 14α-demethylase (CYP51) inhibitors. The photocaged triazoles completely shielded the CYP51 inhibition. The content of ergosterol in fungi before photoactivation and after photoactivation was 4.4% and 83.7%, respectively. Importantly, the shielded antifungal activity (MIC80 ≥ 64 μg/mL) could be efficiently recovered (MIC80 = 0.5-8 μg/mL) by light irradiation. The new chemical tools enable optical control of fungal growth arrest, morphological conversion and biofilm formation. The ability for high-precision antifungal treatment was validated by in vivo models. The light-activated compound A1 was comparable to fluconazole in prolonging survival in Galleria mellonella larvae with a median survival of 14 days and reducing fungal burden in the mouse skin infection model. Overall, this study paves the way for precise regulation of antifungal therapy with improved efficacy and safety.

The stepwise selection for ketoconazole resistance induces upregulation of C14-demethylase (CYP51) in Leishmania amazonensis

Ketoconazole is a clinically safe antifungal agent that also inhibits the growth of Leishmania spp. A study was undertaken to determine whether Leishmania parasites are prone to becoming resistant to ketoconazole by upregulating C14-demethylase after stepwise pharmacological pressure. Leishmania amazonensis promastigotes [inhibitory concentration (IC)50 = 2 µM] were subjected to stepwise selection with ketoconazole and two resistant lines were obtained, La8 (IC50 = 8 µM) and La10 (IC50 = 10 µM). As a result, we found that the resistance level was directly proportional to the C14-demethylase mRNA expression level; we also observed that expression levels were six and 12 times higher in La8 and La10, respectively. This is the first demonstration that L. amazonensis can up-regulate C14-demethylase in response to drug pressure and this report contributes to the understanding of the mechanisms of parasite resistance.

Identification and Functional Characterization of a Cryptococcus neoformans UPC2 Homolog

Azoles are currently the most widely used class of antifungal drugs clinically, and are effective for treating fungal infections. Target site of azoles is ergosterol biosynthesis in fungal cell membrane, which is absent in the mammalian host. However, the development of resistance to azole treatments in the fungal pathogen has become a significant challenge. Here, we report the identification and functional characterization of a UPC2 homolog in the human pathogen Cryptococcus neoformans. UPC2 plays roles in ergosterol biosynthesis, which is also affected by the availability of iron in Saccharomyces cerevisiae and Candida albicans. C. neoformans mutants lacking UPC2 were constructed, and a number of phenotypic characteristics, including antifungal susceptibility and iron utilization, were analyzed. No differences were found between the mutant phenotypes and wild type, suggesting that the role of C. neoformans UPC2 homolog may be different from those in S. cerevisiae and C. albicans, and that the gene may have a yet unknown function.